180 USMLE Road Map: Biochemistry N c. Silent mutations, which often occur in the 3′ base of a codon, do not alter codon specificity, so there is no effect on the protein’s sequence. B. Insertions and deletions may cause production of altered proteins that have ei- ther subtle or drastic functional changes. 1. If the DNA sequence is altered by deletion or insertion of 3n nucleotides, then the mutant protein will lack or gain n amino acid(s), but its sequence will oth- erwise be normal. 2. Insertion or deletion of a number of nucleotides that is not divisible by three will cause a frameshift such that different, garbled protein sequence will be synthesized downstream of the mutation. Single-base substitutions Insertions and deletions Splicing error Silent AUG-UCA-AGA-CCG-AAU-GGC-UAC-UUC-GAU-CUA-AUA Met-Ser-Arg-Pro-Asn-Gly-Tyr-Phe-Asp-Leu-Ile Normal AUG-UCA-AGA-CCG-AAC-GGC-UAC-UUC-GAU-CUA-AUA Met-Ser-Arg-Pro-Asn-Gly-Tyr-Phe-Asp-Leu-Ile Insert (+1 nt) Garbled protein sequence AUG-UGC-AAG-ACC-GAA-CGG-CUA-CUU-CGA-UCU-AAU-A Met-Cys-Lys-Thr-Glu-Arg-Leu-Leu-Arg-Ser-Asn- Nonsense Truncated protein AUG-UCA-AGA-CCG-AAC-GGC-UAG-UUC-GAU-CUA-AUA Met-Ser-Arg-Pro-Asn-Gly-STOP Delete (—1 nt) Garbled sequence, truncated protein AUG-UCA-AGA-cCGA-ACG-GCU-ACU-UCG-AUC-UAA-UA Met-Ser-Arg Arg-Thr-Ala-Thr-Ser-Ile-STOP Garbled sequence AUG-UCA-AGA-CCG-AAC-GGC-AGA-AGC-UGU-GUC-AAA Met-Ser-Arg-Pro-Asn-Gly-Arg-Ser-Cys-Val-Lys Missense AUG-UCA-AGA-CCG-AAC-GUC-UAC-UUC-GAU-CUA-AUA Met-Ser-Arg-Pro-Asn-Val-Tyr-Phe-Asp-Leu-Ile Figure 12–6. Mutations of gene sequences that may affect protein function and cause disease. Chapter 12: Gene Expression 181 N C. Splicing errors alter the critical sequence around an intron-exon splice junction. 1. This may be caused by single-base substitutions, insertions, or deletions. 2. Creation of an abnormal splicing site or destruction of the normal site may re- sult in incorporation of an intron into a “finished” mRNA. 3. Translation of the intron region of the mutant mRNA produces a garbled pro- tein sequence until an in-frame stop codon causes termination of the trun- cated, mutant protein. CLINICAL PROBLEMS A 9-year-old boy is referred for evaluation of his hearing. A note from his school principal explains that he is inattentive in class. Initial physical examination indicates that he is at the 10th percentile for height, has coarse facial features, and is somewhat macrocephalic; however, the remainder of the examination is within normal limits. Audiometry results confirm partial bilateral deafness, which is sensorineural in etiology. An IQ examination shows that he is in the 60th percentile for intelligence. Family history of mucopolysaccha- ridoses prompts specialty testing, which indicates elevated levels of dermatan sulfate and heparan sulfate in both a skin biopsy and urine sample. 1. Biochemical analysis of a skin biopsy from this patient would most likely indicate a de- ficiency of which of the following enzymes? A. β-Galactosidase B. α-L-Iduronidase C. Iduronate sulfatase D. N-Acetylgalactosamine sulfatase E. β-Glucuronidase The sickled shape of erythrocytes in patients with sickle cell anemia occurs because of the tendency for HbS to polymerize. HbS differs from HbA by substitution of a solvent- exposed glutamate by valine in β-globin, which forms a “sticky” patch that promotes aggregation and polymerization of the protein. 2. The genetic change that produced the mutant hemoglobin in sickle cell anemia can be classified as which type of mutation? A. Silent B. Missense C. Nonsense D. Insertion E. Deletion Infections by the ulcer-causing bacterium Helicobacter pylori can be treated effectively with a prolonged course of doxycycline or another of the tetracycline family of antibiotics, po- tent inhibitors of prokaryotic protein synthesis. 182 USMLE Road Map: Biochemistry N 3. Which of the following explains why tetracycline is selective for prokaryotes and mini- mally toxic to humans? A. It is ineffective against the 70S ribosomes. B. It is ineffective against the mitochondrial ribosomes. C. It only inhibits prokaryotic peptidyl transferase. D. It cannot pass across eukaryotic membranes. E. It blocks the A site only of prokaryotic ribosomes. Some patients with familial hypercholesterolemia produce a truncated form of the LDL receptor, termed the “Lebanese” allele, which lacks three of the five domains of the protein and causes it to be retained in the endoplasmic reticulum. Analysis of the mutant gene in- dicated that the sequence of the protein was normal up to the point where it terminated. 4. The genetic change that produced the mutant LDL receptor in these cases can be clas- sified as which type of mutation? A. Silent B. Missense C. Nonsense D. Insertion E. Deletion A 2-year-old boy in whom Down syndrome was diagnosed when he was an infant comes in for a check-up. Although he is developmentally delayed indicating potential mental re- tardation, he is exhibiting some clinical features that are inconsistent with Down syn- drome. These features include coarse facial features, small stature, radiographic evidence of kyphoscoliosis, widening of the ribs, and malformed vertebrae. 5. Microscopic examination of skin or muscle biopsy specimens from this patient would be likely to reveal dense inclusions corresponding with which organelles? A. Mitochondria B. Nuclei C. Golgi apparatus D. Lysosomes E. Peroxisomes A 17-year-old woman with cystic fibrosis is evaluated for knee pain. On review of systems, she also notes persistent bleeding from cuts in her skin and bleeding of her gums after brushing her teeth. Physical examination is remarkable for an obviously swollen right knee that is tender with limited range of motion. Fluid drained from the knee is bloody (hemarthrosis). Her complete blood count is normal, but prothrombin time is elevated. 6. This patient appears to be suffering from a deficiency of which of the following vita- mins? A. Vitamin A B. Vitamin B 12 C. Vitamin C D. Vitamin D E. Vitamin K ANSWERS 1. The answer is C. This patient’s clinical presentation is consistent with one of the mu- copolysaccharidoses, but it can be difficult to determine which type given the wide variability of expression of these disorders. One clue is provided by the hearing loss, a characteristic feature of MPS-II, Hunter syndrome. In addition, his above-average in- telligence for his age group and the absence of scoliosis distinguish MPS-II from MPS I, the Hurler-Scheie syndromes. The latter are characterized by mental retardation to varying degrees. The patient appears to have a severe form of Hunter syndrome, so the cells of his tissues should be deficient in the lysosomal enzyme iduronate sulfatase. 2. The answer is B. A missense mutation results from a change in codon specificity from one amino acid to another. This alters the protein sequence and may affect the pro- tein’s structure and function. By definition, substitution of valine for glutamic acid in the β-globin molecule represents a missense mutation at the level of the gene. Sickle cell anemia illustrates how important even a single amino acid in a large protein can be to the function of the protein and the physiology of the cell. However, it is more com- mon to find that missense mutations have less dramatic effects than in this case. 3. The answer is D. Tetracycline antibiotics operate by blocking the aminoacyl binding site of 30S ribosomes found both in prokayotes and in the mitochondria of eukaryotes. However, the drug may be used as a selective antibiotic with minimal toxicity to pa- tients because it cannot pass through the plasma membranes of human cells. If it could do so, the drug would be cytotoxic because it would interfere with mitochondrial func- tion by inhibiting protein synthesis on the 70S ribosomes of the organelles. 4. The answer is C. Production of a truncated protein indicates that a mutation has oc- curred, but this phenomenon may have arisen from a frameshift mutation (insertion or deletion) or by a nonsense mutation. The most likely possibility is a nonsense mutation because sequence analysis of the truncated protein showed that it had normal (wild- type) sequence. Insertion and deletion events often produce a stretch of garbled or ab- normal protein sequence at the C-terminal end of the truncated protein arising from out-of-frame translation of the mRNA downstream of the mutation until a stop codon is encountered. 5. The answer is D. As this patient ages, a variety of skeletal defects and short stature that are consistent with a lysosomal storage disease (mucolipidosis), either I-cell disease or pseudo-Hurler polydystrophy, are developing. Both diseases arise from a deficiency of an enzyme involved in synthesis of the Man-6-P marker on lysosomal enzymes. Such “misaddressed” proteins are secreted rather than trafficked to the lysosomes. The degradative function of lysosomes is impaired as a result and the organelles tend to ac- cumulate waste products (hence, the term “storage disease”). It is these inclusion bodies or dense structures that would be visible by microscopic examination of the patient’s cells in a biopsy specimen. Chapter 12: Gene Expression 183 N 6. The answer is E. The patient’s symptoms and prolonged prothrombin time suggest that she has a mild coagulation disorder potentially due to vitamin K deficiency. Sev- eral coagulation factors including prothrombin require carboxylation on glutamic acid residues for optimal function. These proteins are carboxylated in vitamin K–dependent reactions. Vitamin K deficiency may occur in people suffering from cystic fibrosis, which causes gastrointestinal complications due to pancreatic insufficiency. Secretion of pancreatic enzymes such as lipase, which releases fatty acids from triglycerides to fa- cilitate absorption from the gut, is impaired in cystic fibrosis patients. This fat malab- sorption condition has manifested itself in this patient’s case as a deficiency in the fat-soluble vitamin K. Although bleeding gums are one characteristic of scurvy, other manifestations of vitamin C deficiency, eg, loose teeth, are absent in this case. 184 USMLE Road Map: Biochemistry N I. Overview of Mendelian Inheritance A. A gene is defined as a unit of DNA that encodes an RNA product. 1. The RNA product may encode transfer RNAs (tRNAs), ribosomal RNAs (rRNAs), or small nuclear RNAs (snRNAs) that have end point functions in the cell. 2. If the RNA product is a messenger RNA (mRNA), then it must be translated into a protein to complete expression of the gene. 3. Variants of a gene that differ in DNA sequence among individuals in the popu- lation are called alleles. a. The single most prevalent version of the gene in the population is referred to as the wild-type (“normal”) allele. b. If there is more than one common version of the gene in the population, these are called polymorphisms. c. Mutant alleles are versions of the gene that differ in sequence from the wild- type allele and that produce defective products. d. The chromosomal location of a gene is its locus. B. The set of alleles that make up the genetic composition of a person is called the genotype, which may refer either to all genes or to a specific gene or locus. 1. The diploid content of human cells is 46 chromosomes—22 autosomal pairs and 2 sex chromosomes (XX in females, XY in males). 2. For genes located on the autosomes, the genotype at a locus is formed from two alleles. 3. Each parent contributes one allele through random segregation of chromo- somes during meiosis. 4. If both alleles at a locus are identical, the person is said to be homozygous for that gene. 5. In the case where the two alleles are different, the person is heterozygous for that gene. 6. Since males have only one X chromosome, they usually have only a single allele and are thus hemizygous for all X-linked genes. C. The measurable expression of the genotype as a molecular, clinical, or biochemical trait is the phenotype. D. Pedigree analysis evaluates transmission of a single-gene disorder within a family or kindred (Figure 13–1). N CHAPTER 13 CHAPTER 13 HUMAN GENETICS 185 Copyright © 2007 by The McGraw-Hill Companies, Inc. Click here for terms of use. II. Modes of Inheritance in Single-Gene Disorders A. In autosomal recessive inheritance, the condition is expressed only in persons who have two copies of (ie, are homozygous for) the mutant allele (Figure 13–2). 1. Autosomal recessive inheritance is often observed with enzyme deficiencies, where heterozygotes express 50% of normal activity. a. However, 50% of normal enzyme activity in these cases permits normal physiologic function because expression of enzyme from the normal allele is sufficient to provide for the needs of the cell. b. This phenomenon is often called the margin of safety effect. 2. Both parents of an affected person for an autosomal recessive disorder must have one normal and one mutant allele, making them obligate carriers barring very rare new mutations. 3. The likelihood of a person being homozygous for an autosomal recessive trait increases in consanguineous matings because of the existence of a common ancestor. 4. Rare autosomal recessive diseases also occur with high frequency among genet- ically isolated populations due to inbreeding. TAYS-SACHS DISEASE IN A GENETICALLY ISOLATED POPULATION • The biochemical defect in Tay-Sachs disease is an inherited deficiency of ␤-hexosaminidase, a lysoso- mal enzyme responsible for hydrolysis of GM 2 ganglioside, which accumulates abnormally in the lyso- somes. • Children with Tay-Sachs disease exhibit hypotonia (poor muscle tone) and progressive neurologic symptoms, including blindness and mental retardation. 186 USMLE Road Map: Biochemistry N Male Female Affected Unaffected Deceased Obligate carrier Marriage or mating Consanguinity Proband 1234 12345 12 3 123 4567 I II III IV , , , , Figure 13–1. Definitions of symbols used to evaluate inheritance patterns for pedigree analysis and relationships within kindreds. Generations are assigned Roman numerals and individuals within each generation are indicated by Arabic numerals. The arrow points at the proband, the person in whom the genetic disorder was first diagnosed. CLINICAL CORRELATION • Most patients are diagnosed at 5–6 months and do not live beyond their second year. • This autosomal recessive disease occurs in Ashkenazi Jews, the Pennsylvania Amish, and several other populations with an incidence of 1 in 3600, 100 times higher than the overall population; the car- rier frequency in these populations is about 3%. 5. Pedigree charts for an autosomal recessive disorder may show the following: a. The disease phenotype is expressed by siblings but not by their parents or offspring. b. Equal occurrence in males and females. c. Recurrence risk for each sibling is 25%. d. Possible consanguinity. Chapter 13: Human Genetics 187 N A I II III IV B I II III IV C I II III IV Figure 13–2. Pedigrees illustrating autosomal inheritance patterns. Recessive in- heritance is shown in pedigrees A and B. Note that consanguinity in pedigree B re- inforces the hypothesis of an autosomal recessive disorder. Dominant inheritance is shown in pedigree C, in which every affected person has an affected parent. B. In autosomal dominant inheritance, the condition is expressed even if a single mutant allele is present, ie, in the heterozygous state (Figure 13–2). 1. Following are at least four possible situations by which having one normal copy of a gene is insufficient to prevent disease, leading to a dominant phenotype (Table 13–1): a. When the presence of 50% normal activity (ordinarily the margin of safety) is not generous enough to allow normal physiologic function, a condition called haploinsufficiency. b. When the defective allele produces a malfunctioning protein product that binds to and interferes with function of the normal gene product—the dominant negative effect. c. When the mutant protein has an enhanced function that overrides normal controls or is cytotoxic. d. When the phenotype appears as dominant inheritance even though the ac- tual allele is recessive at the level of function in individual cells. 2. The homozygous mutant state usually produces a more severe clinical con- dition than the heterozygous condition in autosomal dominant diseases. 3. Pedigree charts for an autosomal dominant disorder may show the following features: a. The disease phenotype appears in all generations, with each affected person having an affected parent. b. There is an equal occurrence in males and females, except in cases when ex- pression of the trait is influenced by the person’s sex (ie, sex-limited). c. Risk of transmission of the mutant allele is 50%, but because there usually are so few persons in a family, there may be deviations from this expectation. d. Potential for some cases to be due to a new mutation, which is more likely for a dominant condition because disease symptoms would be expressed in heterozygotes. 188 USMLE Road Map: Biochemistry N Table 13–1. Molecular phenotypes of autosomal dominant disease. a Molecular Explanation Disease and Gene Haploinsufficiency, or when 50% of normal gene α-Thalassemia trait and the α-globin gene activity is inadequate β-Thalassemia trait and the β-globin gene Dominant negative effect, when the mutant Osteogenesis imperfecta and the collagen 1A protein interferes with function of the normal gene (COL1A1) protein Marfan syndrome and the fibrillin-1 gene (FBN1) Cytotoxic effect due to a dysregulated, mutant Huntington disease and the huntingtin gene (HD) protein Dominant effect at the cellular level of a Retinoblastoma and RB1 recessively inherited loss-of-function mutant Li-Fraumeni syndrome and TP53 of a tumor suppressor gene (see Chapter 14) a These genetic diseases are examples of the various molecular explanations for dominant inheritance. FIBRILLIN DEFECTS IN MARFAN SYNDROME • Marfan syndrome is a connective tissue disorder with manifestations in many organs, but especially the skeleton, blood vessels, eyes, and lungs. • Many tissues, such as lung, blood vessels, and skin, require elasticity for proper function; this prop- erty is fulfilled by the matrix elastic fibers, which are composed of the proteins elastin and fibrillin. – Marfan syndrome arises from a mutation in the gene encoding fibrillin-1 (FBN1). – The pattern of inheritance of Marfan syndrome is autosomal dominant due to the failure of elastic fibers to assemble properly upon interaction of mutant fibrillin with normal elastin. • The disease is usually diagnosed by adolescence, and patients exhibit tall stature and a variety of skeletal deformities, including very long, thin bones of the digits and limbs; flat feet; scoliosis; and breastbone deformation. – Joint hypermobility and a positive wrist/thumb sign are evident. – The upper segment is the distance from the top of the head to the top of the pubic symphysis; the lower segment is the distance from the top of the pubic symphysis to the floor. The upper segment to lower segment ratio in persons with Marfan syndrome is low (< 0.9) because the arms and legs are long relative to the torso. • Characteristic ocular features of Marfan syndrome, such as ectopia lentis (upward lens dislocation in- stead of downward dislocation as in homocystinuria) and myopia, arise from the effects of defective fibrillin in the elastic fibers of the lens. • The major cardiovascular manifestations are mitral valve prolapse and loss of elasticity of the aortic root, which may lead to progressive aneurysm and potentially fatal aortic dissection. C. Most X-linked diseases show a recessive inheritance pattern (Figure 13–3). Chapter 13: Human Genetics 189 N CLINICAL CORRELATION A I II III IV B I II III Figure 13–3. Pedigrees illustrating X-linked recessive (A) and dominant (B) inheri- tance patterns. Note the absence of male-to-male transmission in both pedigrees and the predominance of affected males over females in the X-linked recessive pedigree. [...]... deafness – MERRF (myoclonic epilepsy with ragged red fibers) is characterized by weakness on exertion, ataxia, and associated deafness and is due to mutation of the mitochondrial tRNALys gene – MELAS (mitochondrial encephalomyopathy with lactic acidosis and stroke-like episodes) results from a point mutation in the mitochondrial tRNALeu gene CLINICAL CORRELATION N 192 USMLE Road Map: Biochemistry – LHON... condition from birth as well as macroglossia (enlarged tongue) CLINICAL CORRELATION N 194 USMLE Road Map: Biochemistry • BWS is associated with severe hypoglycemia that may become life-threatening in addition to an enhanced tendency to develop cancers of the liver, kidney, and adrenal glands • The gene for BWS has been mapped to chromosome 11 (11p15), a region encompassing the gene for insulin-like growth... heterozygous females D Occurrence in siblings but not in their parents E Tendency for the disease to skip generations 9 Angelman and Prader-Willi syndromes are related disorders affecting genes on chromosome 15 by which of the following epigenetic mechanisms? N 198 USMLE Road Map: Biochemistry A B C D E Mosaicism Histone acetylation Haploinsufficiency Imprinting Viral infection ANSWERS 1 The answer... Hardy-Weinberg principles based on these same data 2pq = 2(0.02)(0 .98 ) = 0.0 392 or 3 .9% Females Males A a (p) (q) A (p) AA (p2) Aa (pq) a (q) A (pq) aa (q2) Figure 1 3–5 Punnett square showing application of the Hardy-Weinberg Law The allele frequencies p and q are assumed to be equal for males and females within the population N Chapter 13: Human Genetics 195 C The Hardy-Weinberg Law is based on several important... specifically, the patient’s father, maternal grandfather, aunt, uncle, and several cousins are affected 2 Based on these findings, what is the most likely mode of inheritance for the disorder? N 196 USMLE Road Map: Biochemistry A B C D E Autosomal recessive Autosomal dominant X-linked recessive X-linked dominant Mitochondrial 3 The incidence of sickle cell anemia among blacks is 1 in 400 Calculate the frequency...N 190 USMLE Road Map: Biochemistry 1 A distinguishing feature of these diseases is that there can be no male-to-male transmission because the sex of male offspring is determined by contribution of a Y chromosome from... dissociates from the effector and then binds a βγ complex to be ready to undergo a new round of activation N 202 USMLE Road Map: Biochemistry OFF Receptor α β γ GDP G protein Ligand Ligand binding GDP GTP GDP Activated Gα Activated βγ complex ON β α γ GTP Effectors Effectors Figure 1 4–1 Signaling via G protein-coupled receptors Ligand binding to its cellsurface receptor initiates interaction of the... the ovum provides all mitochondria to the fertilized embryo (Figure 1 3–4 ) 3 In these disorders, affected cells usually have a mixture of mitochondria, some with mutant mtDNA and others with wild-type mtDNA, a condition called heteroplasmy N Chapter 13: Human Genetics 191 A I II III IV B 62 I II III 54 43 48 35 48 30 23 29 Figure 1 3–4 Pedigrees illustrating inheritance of (A) a mitochondrial disorder... answer is B The incidence of the disease among blacks, 1/400, can be used to determine q and p: q = 1 = 1/20 = 0.05 and p = 1- q = 1- 0.05 = 0 .95 A 400 The frequency of heterozygotes is then calculated using Hardy-Weinberg Law: Frequency =2 pq = 2(0 .95 )(0.05) = 0. 095 or ∼1 in 11 4 The answer is A The pedigree demonstrates that the age of disease onset decreases in each successive generation, suggesting... phenomena? A Mosaicism B Anticipation C Silencing D Variable expression E Uniparental disomy N Chapter 13: Human Genetics 197 You have assembled a pedigree for the occurrence of hemophilia A in a family of GreekAmerican heritage 1 2 I 1 III 1 1 2 3 2 II 3 4 4 5 6 5 7 6 8 9 7 10 8 11 9 12 13 2 6 Based on these data, what is the most likely mode of inheritance for this disorder? A Autosomal recessive B . principles based on these same data. 2pq = 2(0.02)(0 .98 ) = 0.0 392 or 3 .9% q = A 1 2500 = 20.0004 = 0.02, and: p = 1- q = 1- 0.02 = 0 .98 194 USMLE Road Map: Biochemistry N Males A (p) A (p) a (q) a (q) AA (p 2 ) Aa (pq) A (pq) aa (q 2 ) Females Figure. = A 1 400 = 1/20 = 0.05 and p = 1- q = 1- 0.05 = 0 .95 P = ¢ 1 2 ≤¢ 1 2 ≤¢ 2 3 ≤ A 1 25,000 = ¢ 1 6 ≤¢ 1 500 ≤ = 1 3000 198 USMLE Road Map: Biochemistry N or other genes. Anticipation would have. 1 3–4 ). 3. In these disorders, affected cells usually have a mixture of mitochondria, some with mutant mtDNA and others with wild-type mtDNA, a condition called heteroplasmy. 190 USMLE Road Map: